A large body of data has been accumulated that implicates a family of receptors, the selectins (LEC-CAMs), in certain diseases including cancer, autoimmunity, and in the inflammatory response. The three known members of this family, L-Selectin (LECAM-1, LAM-1, gp90MEL), E-Selectin (LECAM-2, ELAM-1) and P-Selectin (LECAM-3, GMP-140, PADGEM), each contain a domain with homology to the calcium-dependent lectins (C-lectins), an EGF-like domain, and several complement binding protein-like domains (Bevilacqua et al., Science (1989) 243:1160-1165; Johnston et al., Cell (1989) 56:1033-1044; Lasky et al., Cell (1989)56:1045-1055; Tedder et al., J. Exp. Med. (1989) 170.:123-133, Dasgupta et al., Exp. Opin. Invest. Drugs (1994) 3(7):709). It has been proposed that the selectins bind to particular ligands and that this binding accounts for their biological activity. Thus, drugs that interfere with or prevent binding of the ligands to the selectins will be useful medicaments for treating a variety of diseases.
For instance, adhesion of circulating neutrophils to stimulated vascular endothelium is a primary event of the inflammatory response. Recently, Buerke et al. have demonstrated the important role of selectins in inflammatory states such as ischemia-reperfusion injury in cats (Buerke, M. et al, J. Clin. Invest. (1994) 93:1140). Turunen et al. have demonstrated the role of sLe.sup.x and L-selectin in site-specific lymphocyte extravasation in renal transplants during acute rejection (Turunen, J. P. et al, Eur. J. Immunol. (1994) 24:1130). P-selectin has been shown to be centrally involved particularly as related to acute lung injury. Mulligan et al. have reported strong protective effects using anti-P-selectin antibody in a rodent lung injury model. (Mulligan, M. S. et al., J. Clin. Invest. (1991)90:1600, Mulligan, M. S. et al., Nature (1993) 364:149). A central role of P-selectin in inflammation and thrombosis has been demonstrated by Palabrica et at (Palabrica, T. et al., Nature (1992) 359:843).
E-selectin is particularly interesting of the three selectins because of its transient expression on endothelial cells in response to IL-1 or TNF (Bevilacqua et al., Science (1989) 243:1160). The time course of this induced expression (2-8 h) suggests a role for this receptor in initial neutrophil extravasation in response to infection and injury. Indeed, Gundel et al. have shown that antibody to E-selectin blocks the influx of neutrophils in a primate model of asthma and thus is beneficial for preventing airway obstruction resulting from the inflammatory response. (Gundel R. H. et al., J. Clin. Invest. (1991) 88:1407).
Several different groups have published papers regarding E-selectin ligands. Lowe et al., (1990) demonstrated a positive correlation between E-selectin dependent adhesion of HL-60 cell variants and transfected cell lines, with their expression of the sialyl Lewis x (sLe.sup.x) oligosaccharide, NeuNAc .alpha.-2-3-Gal-.beta.1-4(Fuc .alpha.-1-3)-GlcNAc. By transfecting cells with plasmids containing an .alpha.-(1,3/1,4) fucosyltransferase, they were able to convert non-myeloid COS or CHO lines into xLe.sup.x -positive cells that bind in an E-selectin dependent manner. Walz et al., (1990) were able to inhibit the binding of a E-selectin-lgG chimera to HL-60 cells with a monoclonal antibody directed against sLe.sup.x or by glycoproteins with the sLe.sup.x structure, but could not demonstrate inhibition with CD65 or CD15 antibodies. Both groups concluded that the sLe.sup.x structure is the ligand for E-selectin.
Information regarding the DNA sequences encoding endothelial cell-leukocyte adhesion molecules are disclosed in PCT published application WO90/13300 published Nov. 15, 1990 incorporated herein by reference. The PCT publication cites numerous articles which may be related to endothelial cell-leukocyte adhesion molecules. The PCT publication claims methods of identifying E-selectin ligands, as well as methods of inhibiting adhesion between leukocytes and endothelial cells using such ligands and specifically refers to MILAs which are described as molecules involved in leukocyte adhesion to endothelial cells. Recent publications regarding selectin ligands describe the use of L-selectin as an indicator of neutrophil activation (Butcher et al., U.S. Pat. No. 5,316,913 issued May 31, 1994), and assays for inhibition of leukocyte adhesion (Rosen et al., U.S. Pat. No. 5,318,890 issued Jun. 7, 1994).
As alluded to above, the ligand for E-selectin, sLe.sup.x, consists of at least sialic acid, fucose, and lactose. Lactose consists of galactose and glucose. Sialic acid and fucose are bound to the galactose and glucose moieties of lactose, respectively. Ligands that bind to the other selectins share similar structural features. Considering the obvious medical importance of selectin ligands, significant effort has been, and continues to be expended to identify the critical physical/chemical parameters associated with selectin ligands that enhance, or that are required for their activity (DeFrees, S. A., et al, J. Am. Chem., Soc., (1993) 115:7549). In no small part this effort is being driven by the need to have selectin ligands that are inexpensive to produce (see U.S. Pat. No. 5,296,594 issued Mar. 22, 1994, Allanson, N. M. et al., Tetrahedron Lett., (1993) 34:3945.). It is generally thought that it will be commercially prohibitively expensive to produce naturally occurring sLe.sup.x by either enzymatic or chemical synthesis because of the number of sophisticated reactions involved.